Novel Tetrapeptide Inhibitors of Bacterial Protein Synthesis Produced by a Streptomyces sp.

Abstract

In the course of a microbial product screening aimed at the discovery of novel antibiotics acting on bacterial protein synthesis, a complex of three structurally related tetrapeptides, namely, GE81112 factors A, B, and B1, was isolated from a Streptomyces sp. The screening was based on a cell-free assay of bacterial protein synthesis driven by a model mRNA containing natural initiation signals. In this study we report the production, isolation, and structure determination of these novel, potent and selective inhibitors of cell-free bacterial protein synthesis, which stably bind the 30S ribosomal subunit and inhibit the formation of fMet-puromycin. They did not inhibit translation by yeast ribosomes in vitro. Spectroscopic analyses revealed that they are tetrapeptides constituted by uncommon amino acids. While GE81112 factors A, B, and B1 were effective in inhibiting bacterial protein synthesis in vitro, they were less active against Gram-positive and Gram-negative bacterial cells. Cells grown in minimal medium were more susceptible to the compounds than those grown in rich medium, and this is most likely due to competition or regulation by medium components during peptide uptake. The novelty of the chemical structure and of the specific mode of action on the initiation phase of bacterial protein synthesis makes GE81112 a unique scaffold for designing new drugs.

Antibiotic discovery in the twenty-first century: current trends and future perspectives

“…33 The compound was highly effective against a few Grampositive and Gram-negative strains, if grown in minimal or chemically defined medium, suggesting active uptake by the cells. 34 The above examples illustrate how different approaches can lead to novel antibiotic classes. Usually, when unexploited microbial diversity is accessed, there is no need for specific, high-sensitivity assays.…”

Insights into an Unusual Nonribosomal Peptide Synthetase Biosynthesis

“…It should be noted that the major congeners of the GE81112 complex produced by Streptomyces S. sp. L-49973 are A and B, 5 indicating that in vivo O-HPA or a precursor is the preferred substrate for the NRPS.…”

“…Chemical Analyses-GE81112 was purified and hydrolyzed as described by Brandi et al (5). Dehalogenation of GE81112 was carried out under H 2 at atmospheric pressure and room temperature in 10% acetic acid, with 10% palladium/carbon as catalyst.…”

“…A sample of (2R,3R)-3-hydroxypipecolic acid was kindly provided from Prof. Jieping Zhu (CNRS, Gif-sur-Yvette, France). Chiral GC-MS analyses were performed on the methyl esters, and trifluoroacetyl derivatives were analyzed using a Finnigan TSQ700 triple stage quadrupole mass spectrometer interfaced with a Varian 3400 gas chromatographer (5 …”

“…1). Extensive NMR and MS studies revealed the tetrapeptides to comprise hydroxypipecolic and hydroxypentanoic acids, an (amino)histidine, and a hydroxychlorohistidine (5). A retro-biosynthetic analysis of the GE81112 core structure suggests a NRPS 2 origin with additional tailoring steps occurring at some point during assembly.…”